Download Project Description_YL

Project: Cellular microRNA
profilings during the infections
of the hepatitis C virus and the
influenza A virus
Yu Li
Postdoc Fellow
Katze Lab
Projects
1.
Cellular microRNA profilings in the Hepatitis C
virus infected human liver tissues
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2.
HCV positive liver biopsies from transplant patients
HCV negative liver biopsies from transplant patients
Cellular microRNA profilings in the Influenza virus
infected human cell line and mouse lung tissues.
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Viruses:
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H1N1 (human flu): 1918 (High path) and K173 (Low path)
H5N1 (Avian flu): HK483 (High path) and HK486 (Low path)
Mock virus
In vitro study- Human epithelial cells A549
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H1N1, H5N1 influenza flu viruses and mock virus infections
In vivo study- Mice lungs from a previously published study
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H1N1 only- 1918(High path), Texas (Low path) and Mock virus
MicroRNA
• MicroRNA is a group of noncoding small RNA
molecules with ~22 nt in length.
– 847 are registered human microRNA in Sanger
database V10 as of Jun 2008
• MicroRNAs play a central role in regulating
cellular gene expression at posttranscriptional
levels.
– On average, a microRNA regulates the expression of
over 200 cellular genes or microRNA targets
– A microRNA targets can be regulated by multiple
microRNAs
Repression of Gene Expression by MicroRNA
RISC
microRNA
Seed region from 2-8
Target mRNA
Cap
3’ UTR
3’ UTR
(A)n
ORF
Cap
Partial match
(A)n
ORF
Near perfect match
3’ UTR
Cap
ORF
(A)n
Cap
(A)n
Translation inhibition
mRNA cleavage
Decreased protein abundance
Experimental Design
A549 cells infected with
high-path virus and low-path influenza virus
(total RNA)
Flu-associated
mRNA expression profile
Flu-associated
microRNA expression profile
(Gene expression arrays)
(MicroRNA arrays)
microRNA
target prediction
Inverse correlation of flu-associated
microRNA and gene expressions
Current status- The HCV-microRNA project
• MicroRNA expression profilings:
– 22 HCV positive liver biopsies (20 finished & 2 will be finished this
week)
– 6 HCV negative liver biopsies (will be finished this week)
– All microRNAs arrays are single channel arrays
• Gene expression profilings:
– 22 HCV positive liver biopsies. (Finished by NIDA)
– All gene expression arrays are duel channel arrays (pooled mock as the
reference)
• MicroRNA target prediction in the human
genome
– Xinxia used “perfect-match” for predicting microRNA
targets that can be repressed by a microRNA on the
mRNA level. (Finished)
Current status- The Flu-microRNA project
• MicroRNA expression profilings:
– H1N1 and H5N1 infected A549 cells (the majority are finished, 2
more slides to finish )
– H1N1 infected mouse lungs (will be finished in 2 weeks)
• Gene expression profilings:
– H1N1, H5N1 and mock infected A549 cells (finished)
– H1N1 and mock infected mouse lung (Kash paper 2006)
– We are not only focused on the difference between high path and low path
flu virus infections but also focused on the difference between flu and
mock.
• MicroRNA target prediction in the human and mouse
genomes
– Xinxia used “perfect-match” for predicting microRNA targets that
can be repressed by a microRNA on the mRNA level. (Finished)
Next steps
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Establish the correlation between the microRNA expression changes and the
gene expression changes:
– 2 independent analyses:
• From the end of microRNA profilings:
– Make the prediction of gene expression changes from microRNA expression changes
• From the end of gene expression profilings:
– Make the prediction of microRNA expression changes from gene expression changes
– Both analyses will help to find the pathways that associated with HCV or
influenza infections and microRNA expression changes.
– The common pathways found by both analyses have higher probability to be
real and would be use to make biological stories.
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Taqman validation of the both microRNA and gene expression changes if the
material is still available:
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Draft a manuscript: